The Notch signaling pathway is a universally conserved signal transduction system. It is involved in cell fate determination during development including embryonic pattern formation and post-embryonic tissue maintenance. In addition, Notch signaling has been identified as a critical factor in the maintenance of hematopoietic stem cells.
The mammalian Notch receptor family includes four members, Notch1, Notch2, Notch3 and Notch4. Notch receptors are large single-pass type I transmembrane proteins with several conserved structural motifs. The extracellular domain contains a variable number of epidermal growth factor (EGF)-like repeats involved in ligand binding and three cysteine-rich LIN-12/Notch repeats (LNRs) involved in Notch heterodimerization. The intracellular domain contains a RAM23 motif involved in binding Notch downstream signaling proteins, 7 cdc10/ankyrin repeats also involved in mediating downstream signaling and a PEST domain involved in Notch protein degradation.
Mammalian Notch ligands include Delta-like 1 (DLL1), Delta-like 3 (DLL3), Delta-like 4 (DLL4), Jagged1 and Jagged2. Similar to Notch receptors, Notch ligands are type I transmembrane proteins with several conserved structural motifs. Extracellular motifs common to all Notch ligands include a single Delta/Serrate/Lag-2 (DSL) domain involved in receptor binding, as well as a variable number of EGF-like repeats that may be involved in stabilizing receptor binding. The extracellular domain of Jagged proteins contains a cysteine-rich region which has partial homology to the von Willebrand factor type C domain and is likely involved in ligand dimerization. This motif is not present in DLL family members. (Leong et al., 2006, Blood, 107:2223-2233).
The extracellular domain of a Notch receptor interacts with the extracellular domain of a Notch ligand, typically on adjacent cells, resulting in two proteolytic cleavages of the Notch receptor. One extracellular cleavage is mediated by an ADAM (A Disintegrin And Metallopeptidase) protease and a second cleavage within the transmembrane domain is mediated by the gamma secretase complex. This latter cleavage generates the Notch intracellular domain (ICD), which translocates to the nucleus where it activates the CBF1, Suppressor of Hairless, Lag-2 (CSL) family of transcription factors as the major downstream effectors to increase transcription of nuclear basic helix-loop-helix transcription factors of the Hairy/Enhancer of Split (HES) family. (Artavanis et al., 1999, Science, 284:770; Brennan and Brown, 2003, Breast Cancer Res., 5:69; Iso et al., 2003, Arterioscler. Thromb. Vase. Biol., 23:543).
The Notch pathway has been linked to the pathogenesis of both hematologic and solid tumors and cancers. Numerous cellular functions and microenvironmental cues associated with tumorigenesis have been shown to be modulated by Notch pathway signaling, including cell proliferation, apoptosis, adhesion, and angiogenesis. (Leong et al., 2006, Blood, 107:2223-2233). In addition, Notch receptors and/or Notch ligands have been shown to play potential oncogenic roles in a number of human cancers, including acute myelogenous leukemia, B cell chronic lymphocytic leukemia, Hodgkin lymphoma, multiple myeloma, T-cell acute lymphoblastic leukemia, brain cancer, breast cancer, cervical cancer, colon cancer, lung cancer, pancreatic cancer, prostate cancer and skin cancer. (Leong et al., 2006, Blood, 107:2223-2233).
The Notch1 gene in humans was first identified in a subset of T-cell acute lymphoblastic leukemias as a translocated locus resulting in activation of the Notch pathway (Ellisen et al., 1991, Cell, 66:649-61). More recently, it has been shown that more than 50% of human T-cell acute lymphoblastic leukemias have activating mutations that involve the extracellular heterodimerization domain and/or the C-terminal PEST domain of Notch1 (Weng et al., 2004, Science, 306:269-271; Pear & Aster, 2004, Curr. Opin. Hematol., 11:416-33). Constitutive activation of Notch1 signaling in T-cells in mouse models similarly generates T-cell lymphomas suggesting a causative role (Robey et al., 1996, Cell, 87:483-92; Pear et al., 1996, J. Exp. Med., 183:2283-91; Yan et al., 2001, Blood, 98:3793-9; Bellavia et al., 2000, EMBO J. 19:3337-48). Retrovirally-activated Notch2 has been implicated in thymic lymphoma induced by feline leukemia virus (Rohn et al., 1996, J. Virology, 70:8071-8080). Human T-cell acute lymphoblastic leukemia samples have been shown to express Notch3 and its target gene HES-1, which were not expressed in normal peripheral T-cells nor in non-T-cell leukemias (Bellavia et al., 2002, PNAS, 99:3788-3793). Thus, the Notch pathway has been identified as a potential target for therapeutic intervention in several hematologic cancers.
Anti-Notch antibodies and their possible use as anti-cancer therapeutics have been reported. See, e.g., U.S. Patent Application Publication Nos. 2008/0131434 and 2009/0081238, each of which is incorporated by reference herein in its entirety. See also International Publication Nos. WO 2008/057144, WO 2008/076960, WO 2008/150525, WO 2010/005566 and WO 2010/005567; each of which is incorporated by reference herein in its entirety.